Enhanced methamphetamine metabolism in rhesus macaque as compared with human: an analysis using a novel method of liquid chromatography with tandem mass spectrometry, kinetic study, and substrate docking.

Methamphetamine (MA), which remains one of the widely used drugs of abuse, is metabolized by the cytochrome P450 (CYP) family of enzymes in humans. However, metabolism of methamphetamine in macaques is poorly understood. Therefore, we first developed and validated a very sensitive LC-MS/MS method using solid phase extraction of rhesus plasma with a lower limit of quantitation at 1.09 ng/mL for MA and its metabolites, 4-hydroxy methamphetamine (4-OHMA), amphetamine (AM), 4-OH amphetamine (4-OHAM), and norephedrine. We then analyzed plasma samples of MA-treated rhesus, which showed >10-fold higher concentrations of AM (~29 ng/mL) and 4-OHAM (~28 ng/mL) than MA (~2 ng/mL). Since the plasma levels of MA metabolites in rhesus were much higher than in human samples, we examined MA metabolism in human and rhesus microsomes. Interestingly, the results showed that AM and 4-OHAM were formed more rapidly and the catalytic efficiency (Vmax/Km) for the formation of AM was ~8-fold higher in rhesus than in human microsomes. We further examined the differences in these kinetic characteristics using three selective inhibitors of each human CYP2D6 and CYP3A4 enzymes. The results showed that each of these inhibitors inhibited both d- and l-MA metabolism by 20-60% in human microsomes, but not in rhesus microsomes. The differences between human and rhesus CYP2D6 and CYP3A4 enzymes were further assessed by docking studies for both d-and l-MA. In conclusion, our results demonstrated an enhanced MA metabolism in rhesus compared to human, which is likely to be caused by differences in MA-metabolizing CYP-enzymes between these species.